Whole exome sequencing identifies causative mutations in the majority of consanguineous or familial cases with childhood-onset increased renal echogenicity
2015; Elsevier BV; Volume: 89; Issue: 2 Linguagem: Inglês
10.1038/ki.2015.317
ISSN1523-1755
AutoresDaniela A. Braun, Markus Schueler, Jan Halbritter, Heon Yung Gee, Jonathan D. Porath, Jennifer A. Lawson, Rannar Airik, Shirlee Shril, Susan J. Allen, Deborah R. Stein, Adila Al Kindy, Bodo B. Beck, Nurcan Cengız, Khemchand N Moorani, Fatih Özaltın, Seema Hashmi, John A. Sayer, Detlef Böckenhauer, Neveen A. Soliman, Edgar A. Otto, Richard P. Lifton, Friedhelm Hildebrandt,
Tópico(s)Genomics and Rare Diseases
ResumoChronically increased echogenicity on renal ultrasound is a sensitive early finding of chronic kidney disease that can be detected before manifestation of other symptoms. Increased echogenicity, however, is not specific for a certain etiology of chronic kidney disease. Here, we performed whole exome sequencing in 79 consanguineous or familial cases of suspected nephronophthisis in order to determine the underlying molecular disease cause. In 50 cases, there was a causative mutation in a known monogenic disease gene. In 32 of these cases whole exome sequencing confirmed the diagnosis of a nephronophthisis-related ciliopathy. In 8 cases it revealed the diagnosis of a renal tubulopathy. The remaining 10 cases were identified as Alport syndrome (4), autosomal-recessive polycystic kidney disease (2), congenital anomalies of the kidney and urinary tract (3), and APECED syndrome (1). In 5 families, in whom mutations in known monogenic genes were excluded, we applied homozygosity mapping for variant filtering and identified 5 novel candidate genes (RBM48, FAM186B, PIAS1, INCENP, and RCOR1) for renal ciliopathies. Thus, whole exome sequencing allows the detection of the causative mutation in 2/3 of affected individuals, thereby presenting the etiologic diagnosis, and allows identification of novel candidate genes. Chronically increased echogenicity on renal ultrasound is a sensitive early finding of chronic kidney disease that can be detected before manifestation of other symptoms. Increased echogenicity, however, is not specific for a certain etiology of chronic kidney disease. Here, we performed whole exome sequencing in 79 consanguineous or familial cases of suspected nephronophthisis in order to determine the underlying molecular disease cause. In 50 cases, there was a causative mutation in a known monogenic disease gene. In 32 of these cases whole exome sequencing confirmed the diagnosis of a nephronophthisis-related ciliopathy. In 8 cases it revealed the diagnosis of a renal tubulopathy. The remaining 10 cases were identified as Alport syndrome (4), autosomal-recessive polycystic kidney disease (2), congenital anomalies of the kidney and urinary tract (3), and APECED syndrome (1). In 5 families, in whom mutations in known monogenic genes were excluded, we applied homozygosity mapping for variant filtering and identified 5 novel candidate genes (RBM48, FAM186B, PIAS1, INCENP, and RCOR1) for renal ciliopathies. Thus, whole exome sequencing allows the detection of the causative mutation in 2/3 of affected individuals, thereby presenting the etiologic diagnosis, and allows identification of novel candidate genes. Renal ultrasound imaging (RUS) represents a simple and broadly accessible tool for the noninvasive early diagnosis of chronic kidney disease (CKD) in children and young adults. Often abnormal findings on RUS are detectable years before the kidney function deteriorates and before other symptoms develop. A typical abnormal finding in early stages of CKD is chronically increased echogenicity on RUS. It is frequently accompanied by loss of cortico-medullary differentiation and renal cysts. Increased echogenicity is easily detected as a degree of echogenicity that is equal to or more pronounced than the echogenicity of the liver. Unfortunately, chronically increased echogenicity is not specific to certain types of kidney disease.1Gee H.Y. Otto E.A. Hurd T.W. et al.Whole-exome resequencing distinguishes cystic kidney diseases from phenocopies in renal ciliopathies.Kidney Int. 2014; 85: 880-887Abstract Full Text Full Text PDF PubMed Scopus (54) Google Scholar, 2Kraus R.A. Gaisie G. Young L.W. Increased renal parenchymal echogenicity: causes in pediatric patients.Radiographics. 1990; 10: 1009-1018Crossref PubMed Scopus (39) Google Scholar, 3Moghazi S. Jones E. Schroepple J. et al.Correlation of renal histopathology with sonographic findings.Kidney Int. 2005; 67: 1515-1520Abstract Full Text Full Text PDF PubMed Scopus (146) Google Scholar Particularly in early stages, in which other symptoms are not yet present, a correct diagnosis can be challenging. In these cases, whole-exome sequencing (WES) provides a novel means of establishing an etiologic diagnosis. By revealing the causative monogenic mutation, it provides affected individuals and their families with an unequivocal, early diagnosis.1Gee H.Y. Otto E.A. Hurd T.W. et al.Whole-exome resequencing distinguishes cystic kidney diseases from phenocopies in renal ciliopathies.Kidney Int. 2014; 85: 880-887Abstract Full Text Full Text PDF PubMed Scopus (54) Google Scholar As a result, a targeted therapeutic regimen can be initiated early if available. Chronically increased echogenicity on RUS is often found in the early stages of nephronophthisis-related ciliopathies (NPHP-RCs). NPHP-RCs represent a group of cystic and fibrotic kidney diseases with an autosomal recessive mode of inheritance that typically progress to end-stage renal failure within the first three decades of life.4Hildebrandt F. Benzing T. Katsanis N. Ciliopathies.N Engl J Med. 2011; 364: 1533-1543Crossref PubMed Scopus (979) Google Scholar, 5Hildebrandt F. Attanasio M. Otto E. Nephronophthisis: disease mechanisms of a ciliopathy.J Am Soc Nephrol. 2009; 20: 23-35Crossref PubMed Scopus (270) Google Scholar Nephronophthisis can present as isolated renal disease (MIM #613550) or together with extrarenal symptoms such as retinal degeneration (Senior Loken syndrome MIM #266900), cerebellar vermis hypoplasia (Joubert syndrome MIM #213300), and hepatic fibrosis. The renal manifestation ranges from severe, early onset cystic kidney disease6Otto E.A. Schermer B. Obara T. et al.Mutations in INVS encoding inversin cause nephronophthisis type 2, linking renal cystic disease to the function of primary cilia and left-right axis determination.Nat Genet. 2003; 34: 413-420Crossref PubMed Scopus (508) Google Scholar to slowly progressive, fibrotic remodeling of the kidney with CKD starting in adolescence.7Olbrich H. Fliegauf M. Hoefele J. et al.Mutations in a novel gene, NPHP3, cause adolescent nephronophthisis, tapeto-retinal degeneration and hepatic fibrosis.Nat Genet. 2003; 34: 455-459Crossref PubMed Scopus (288) Google Scholar Interestingly, the genotype–phenotype correlation in NPHP-RC is dependent on the gene and the specific mutation involved, which can both give rise to a broad phenotypic disease spectrum. NPHP-RCs are a very heterogeneous disease group, as by now mutations in more than 90 genes have been identified as causative for renal ciliopathies in humans.4Hildebrandt F. Benzing T. Katsanis N. Ciliopathies.N Engl J Med. 2011; 364: 1533-1543Crossref PubMed Scopus (979) Google Scholar Mutations in some of these genes are very rare, accounting for only two8Attanasio M. Uhlenhaut N.H. Sousa V.H. et al.Loss of GLIS2 causes nephronophthisis in humans and mice by increased apoptosis and fibrosis.Nat Genet. 2007; 39: 1018-1024Crossref PubMed Scopus (180) Google Scholar or three9Otto E.A. Trapp M.L. Schultheiss U.T. et al.NEK8 mutations affect ciliary and centrosomal localization and may cause nephronophthisis.J Am Soc Nephrol. 2008; 19: 587-592Crossref PubMed Scopus (147) Google Scholar families worldwide. WES with direct inspection of the coding regions of these genes therefore represents the most rational and currently, most cost-effective approach for mutation analysis in these patients.1Gee H.Y. Otto E.A. Hurd T.W. et al.Whole-exome resequencing distinguishes cystic kidney diseases from phenocopies in renal ciliopathies.Kidney Int. 2014; 85: 880-887Abstract Full Text Full Text PDF PubMed Scopus (54) Google Scholar, 10Chaki M. Airik R. Ghosh Amiya K. et al.Exome capture reveals ZNF423 and CEP164 mutations, linking renal ciliopathies to DNA damage response signaling.Cell. 2012; 150: 533-548Abstract Full Text Full Text PDF PubMed Scopus (260) Google Scholar, 11Otto E.A. Hurd T.W. Airik R. et al.Candidate exome capture identifies mutation of SDCCAG8 as the cause of a retinal-renal ciliopathy.Nat Genet. 2010; 42: 840-850Crossref PubMed Scopus (254) Google Scholar, 12Schueler M. Braun D.A. Chandrasekar G. et al.DCDC2 mutations cause a renal-hepatic ciliopathy by disrupting Wnt signaling.Am J Hum Genet. 2015; 96: 81-92Abstract Full Text Full Text PDF PubMed Scopus (68) Google Scholar So far, no more than 13 ciliopathy genes have been systematically studied in a larger patient cohort.13Halbritter J. Diaz K. Chaki M. et al.High-throughput mutation analysis in patients with a nephronophthisis-associated ciliopathy applying multiplexed barcoded array-based PCR amplification and next-generation sequencing.J Med Genet. 2012; 49: 756-767Crossref PubMed Scopus (98) Google Scholar, 14Halbritter J. Porath J.D. Diaz K.A. et al.Identification of 99 novel mutations in a worldwide cohort of 1,056 patients with a nephronophthisis-related ciliopathy.Hum Genet. 2013; 132: 865-884Crossref PubMed Scopus (165) Google Scholar Here, we performed WES combined with homozygosity mapping in an international cohort of 79 families with pediatric onset of CKD and suspected nephronophthisis based on renal ultrasound presentation with chronically increased echogenicity, loss of cortico-medullary differentiation, and/or ≥2 cysts. All individuals were born of consanguineous union, or represented familial cases of CKD, and were therefore at high risk for recessive, monogenic diseases. In summary, we were able to identify a causative mutation in a known monogenic disease gene in 50 families (63.3%). In 32 of these families (64%), WES identified mutations in NPHP-RC genes as the molecular disease cause and confirmed the suspected clinical diagnosis. However, in 18 families (36%), we discovered a molecular diagnosis of a monogenic kidney disease that was not NPHP-RC, specifically renal tubulopathies (n = 8, 16%), Alport syndrome (n = 4, 8%), congenital anomalies of the kidney and urinary tract (CAKUT) (n = 3, 6%), autosomal recessive polycystic kidney disease (ARPKD) (n = 2, 4%), and autoimmune nephropathy (APECED [autoimmune polyendocrinopathy-candidiasis–ectodermal dystrophy] syndrome) (n = 1, 2%). In five consanguineous families, in whom we excluded mutations in known monogenic disease genes, we identified five novel candidate genes for NPHP-RC (RBM48, FAM168B, PIAS1, INCENP, and RCOR1). We performed WES in 79 families with suspected NPHP-RC based on renal ultrasound criteria (chronically increased echogenicity, loss of cortico-medullary differentiation, and/or ≥2 renal cysts). In 50/79 families (63.3%), we identified a mutation in a gene that is known to cause monogenic renal disease when mutated (Figure 1a, Supplementary Tables S1 and S2 online). Thirty-two of the 50 families with established molecular diagnosis after WES (64.0%) harbored a mutation in a known NPHP-RC gene (Figure 1b). Of the 90 genes that are known to cause renal ciliopathies when mutated, which were systematically analyzed in this study, mutations in 18 genes contributed to this result. Mutations in the genes NPHP3, NPHP4, and NPHP5 accounted for the majority of NPHP-RC cases (Table 1, Supplementary Tables S1 and S2 online). In addition to mutations in NPHP-RC genes, we detected causative mutations in monogenic genes of renal tubulopathies (8/50), Alport syndrome (4/50), CAKUT (3/50), and ARPKD (2/50). Furthermore, we established the molecular diagnosis of APECED in one previously undiagnosed individual (Figure 1b, Table 1, Supplementary Tables S1 and S2 online). In 10 individuals, the molecular diagnosis after WES was different from the previous clinical diagnosis. In particular, in one case of previously undiagnosed nephro-pathic cystinosis the molecular finding changed the therapeutic regimen (Supplementary Tables S1 and S2 online).Table 1Diagnostic groups and distribution among specific genes with causative recessive mutations in 50 families with increased echogenicity and/or ≥2 cysts on renal ultrasound in whom the disease-causing mutation was detected by WESDisease groupPatients within the disease groupGene with causative mutation (disease, OMIM#aOnline Mendelian Inheritance in Man (OMIM), http://www.omim.org.)NPHP-RC32/50 (64%)2/50 (4%)NPHP1 (nephronophthisis type, juvenile, #256100)3/50 (6%)NPHP3 (nephronophthisis type 3, #604387)5/50 (10%)NPHP4 (nephronophthisis type 4, #606966)3/50 (6%)IQCB1 (Senior-Loken syndrome type 5, #609254)1/50 (2%)CEP290 (Senior-Loken syndrome 6, #610189)2/50 (4%)SDCCAG8 (Senior-Loken syndrome type 7, #613615)1/50 (2%)TMEM67 (nephronophthisis 11, #613550)2/50 (4%)TTC21B (nephronophthisis 12, #613820)2/50 (4%)WDR19 (nephronophthisis 13, #614377)3/50 (6%)ANKS6 (nephronophthisis 16, #615382)1/50 (2%)TMEM138 (Joubert syndrome 16, #614465)1/50 (2%)TMEM231 (Joubert syndrome 20, #614970)1/50 (2%)ARL6 (Bardet-Biedl syndrome 3, #600151)1/50 (2%)BBS4 (Bardet-Biedl syndrome 4, #615982)1/50 (2%)MKKS (Bardet-Biedl syndrome 6, #605231)1/50 (2%)BBS12 (Bardet-Biedl syndrome 12, #615989)1/50 (2%)DYNC2H1 (short-rib thoracic dysplasia 3, #613091)1/50 (2%)IFT43 (cranioectodermal dysplasia 3, #614099)Renal tubulopathies8/50 (16%)3/50 (6%)CLDN16 (hypomagnesemia 3, renal, #248250)2/50 (4%)CLCNKB (Bartter syndrome, type 3, #607364)1/50 (2%)ATP6V0A4 (renal tubular acidosis, distal, #602722)1/50 (2%)BCS1L (mitochondrial complex III deficiency, #124000)1/50 (2%)CTNS (cystinosis, nephropathic, #219800)Alport syndrome4/50 (8%)2/50 (4%)COL4A3 (Alport syndrome, #203780)1/50 (2%)COL4A4 (Alport syndrome, #203780)1/50 (2%)COL4A5 (Alport syndrome, #301050)CAKUT3/50 (6%)2/50 (4%)HNF1B (renal cysts and diabetes syndrome, #137920)1/50 (2%)FREM2 (Fraser syndrome, #219000)Autosomal recessive polycystic kidney disease2/50 (4%)PKHD1 (polycystic kidney and hepatic disease, #263200)APECED1/50 (2%)AIRE (autoimmune polyendocrinopathy syndrome type I, #240300)APECED, autoimmune polyendocrinopathy-candidiasis–ectodermal dystrophy; CAKUT, congenital anomalies of the kidney and urinary tract; NPHP-RC, nephronophthisis-related ciliopathies.a Online Mendelian Inheritance in Man (OMIM), http://www.omim.org. Open table in a new tab APECED, autoimmune polyendocrinopathy-candidiasis–ectodermal dystrophy; CAKUT, congenital anomalies of the kidney and urinary tract; NPHP-RC, nephronophthisis-related ciliopathies. In one example of a highly inbred family (412 Mb of cumulative homozygosity), we identified homozygous mutations in seven known monogenic disease genes. Four of them are known to cause diseases with renal involvement (Supplementary Table S4 online). The affected child showed a complex phenotype suggestive of a renal ciliopathy, including Caroli's disease with massive cystic dilation of intrahepatic bile ducts, CKD that progressed to end-stage renal failure within the third year of life, post-axial polydactyly, nystagmus, and rod cone dystrophy. In addition to chronically increased echogenicity as the classical symptom of NPHP-RC, the child showed congenital hydronephrosis due to uretero-pelvic junction obstruction that required corrective surgery, as well as renal tubular acidosis type 4 (Supplementary Table S4 online). An older brother, from whom no DNA sample was available for mutation analysis, deceased at 15 months of age due to end-stage renal and liver disease. In addition, he showed post-axial polydactyly and blindness at 5 months of age due to progressive retinal dystrophy. A younger brother presented with Caroli's disease and retinitis pigmentosa. Interestingly, no polydactyly was present, and the renal function was preserved at 3 years of age. The right kidney showed mild pelvicalyceal dilation. When performing WES in 79 families with suspected nephronophthisis based on RUS with childhood-onset chronically increased renal echogenicity and/or ≥2 renal cysts, we detected a causative mutation in a known monogenic disease gene in 38 of 60 consanguineous families (63.3%) (Figure 2). As postulated15Hildebrandt F. Heeringa S.F. Ruschendorf F. et al.A systematic approach to mapping recessive disease genes in individuals from outbred populations.PLoS Genet. 2009; 5: e1000353Crossref PubMed Scopus (133) Google Scholar, 16Ten Kate L.P. Scheffer H. Cornel M.C. et al.Consanguinity sans reproche.Hum Genet. 1991; 86: 295-296Crossref PubMed Scopus (14) Google Scholar in the vast majority of consanguineous families, the causative mutation was present in the homozygous state (37 of 38 families [97.4%]) (Figure 2). The only exception was a family in whom we identified a single heterozygous mutation in the dominant gene HNF1B as causative. In 12 of 19 non-consanguineous families with 2 or more affected children (63.2%), we identified a causative monogenic mutation, showing a comparable percentage of molecularly solved cases in both cohorts. However, as expected in non-consanguineous families,16Ten Kate L.P. Scheffer H. Cornel M.C. et al.Consanguinity sans reproche.Hum Genet. 1991; 86: 295-296Crossref PubMed Scopus (14) Google Scholar only in 3 of 12 families (25.0%) the disease-causing mutation was present in the homozygous state. The remaining 9 of 12 families (75.0%) harbored two compound heterozygous disease-causing mutations (Figure 2). By applying homozygous peak regions as a filter for WES data, we identified five novel candidate genes for monogenic, recessive NPHP-RC (Table 2). In these five consanguineous families, we had excluded mutations in known monogenic disease genes with renal phenotypes through evaluation and direct inspection of the WES data. The five novel candidate genes (RBM48, FAM168B, PIAS1, INCENP, and RCOR) represented the most deleterious alleles within the homozygous regions (Supplementary Figure S2 online) after variant filtering as outlined in Supplementary Figure S1 online. In the example of family A2621 with 181 Mb of cumulative homozygosity (Supplementary Table S5 online), we started with 482,406 variants from normal reference sequence. Excluding common variants (minor allele frequency >1% in healthy control cohorts), synonymous variants, and heterozygous variants reduced the number to 1699. Considering only variants that were positioned within homozygous peak regions reduced the number of remaining variants by 11-fold to 156. Exclusion of artifacts by direct inspection of the sequence alignment left us with three potentially disease-causing variants in this family (Supplementary Table S5 online). Subsequently, these variants were ranked based on their predicted likelihood to be deleterious for the function of the encoded protein following the criteria as outlined in Supplementary Figure S1 online. At the end of the filtering process, the gene INCENP (inner centromere protein) represented the strongest remaining variant in this family. All mutations in novel candidate genes were confirmed by Sanger sequencing in original patient DNA and segregated with the affected status.Table 2Novel candidate genes for NPHP-RC identified in 5 families with increased echogenicity and/or ≥2 cysts on renal ultrasound in whom causative mutations in known monogenic disease genes were excludedFamilyGeneZygosityAccession #c.Positionp.PositionContinuously conserved toPolyphen-2/ MutTast/SIFTExACEVSaGenotypes in European Americans.Clinical diagnosisA2621INCENPHomNM_020238.2C.2403G > Cp.Gln801HisDanio rerio0.96/DC/del0/337/1208260/32/4267NPHPA1833RBM48HomNM_032120.2c.835A>Gp.Thr279AlaCiona intestinalis0.71/DC/del0/5/120562Not presentNPHP+JATDA2275FAM186BHomNM_032130.2c.506-2A>GObligatory splice site mutation0/1/111410Not presentNPHPA2287PIAS 1HomNM_016166.1c.317C>Tp.Ser106LeuDanio rerio0.77/DC/del0/3/120766Not presentNPHPA1239RCOR1HomNM_015156.2c.437-3C>TSplice site mutation0/3/121306Not presentJBTSDC, disease causing (MutationTaster); del, deleterious (SIFT); EVS, exome variant server; ExAC, Exome Aggregation Consortium; Hom, homozygous; JATD, Jeune asphyxiating thoracic dystrophy; JBTS, Joubert syndrome; MutTast, MutationTaster; NPHP, nephronophthisis; NPHP-RC, nephronophthisis-related ciliopathy; SIFT, sorting intolerant from tolerant. Each candidate gene represents the most deleterious mutation within the homozygous peak regions of the respective family.a Genotypes in European Americans. Open table in a new tab DC, disease causing (MutationTaster); del, deleterious (SIFT); EVS, exome variant server; ExAC, Exome Aggregation Consortium; Hom, homozygous; JATD, Jeune asphyxiating thoracic dystrophy; JBTS, Joubert syndrome; MutTast, MutationTaster; NPHP, nephronophthisis; NPHP-RC, nephronophthisis-related ciliopathy; SIFT, sorting intolerant from tolerant. Each candidate gene represents the most deleterious mutation within the homozygous peak regions of the respective family. Twenty-four families (30.4%) remained without a molecular diagnosis after WES. In these families, no convincing variants in known monogenic genes or candidate genes were identified that segregated with the affected status. We performed WES combined with homozygosity mapping in 79 consanguineous or familial cases of childhood-onset chronically increased renal echogenicity or the presence of ≥2 cysts on renal ultrasound. On the basis of renal ultrasound presentation, clinical presentation with childhood-onset of disease, and if available renal histology, an NPHP-RC was suspected as the primary disease cause. We show that, in this patient cohort, WES allows detecting the specific causative mutation in about two-thirds of affected individuals, thereby presenting the etiologic diagnosis. In addition, we identified five potential novel renal ciliopathy genes in consanguineous families in whom mutations in 90 known monogenic NPHP-RC genes had been excluded. Chronically increased echogenicity on renal ultrasound represents a sensitive diagnostic criterion for early stages of CKD in children and young adults. Typically, this results in the suspected diagnosis of nephronophthisis or related ciliopathies. However, we and others1Gee H.Y. Otto E.A. Hurd T.W. et al.Whole-exome resequencing distinguishes cystic kidney diseases from phenocopies in renal ciliopathies.Kidney Int. 2014; 85: 880-887Abstract Full Text Full Text PDF PubMed Scopus (54) Google Scholar, 2Kraus R.A. Gaisie G. Young L.W. Increased renal parenchymal echogenicity: causes in pediatric patients.Radiographics. 1990; 10: 1009-1018Crossref PubMed Scopus (39) Google Scholar, 3Moghazi S. Jones E. Schroepple J. et al.Correlation of renal histopathology with sonographic findings.Kidney Int. 2005; 67: 1515-1520Abstract Full Text Full Text PDF PubMed Scopus (146) Google Scholar have shown that other kidney diseases can phenocopy the presentation of NPHP-RC on renal ultrasound. Here, we study 79 consanguineous or familial cases with suspected NPHP-RC based on renal ultrasound presentation with childhood-onset increased renal echogenicity and/or more than one cyst, and perform WES in order to determine the percentage in which NPHP-RC and various other monogenic, recessive kidney diseases contribute to this phenotypic spectrum (Figure 2). As a second aspect, we aimed to systematically examine all 90 genes that are known monogenic, recessive causes of NPHP-RC or other CKD that can phenocopy the presentation of NPHP-RC in a large cohort of affected individuals. So far, this has not been done for more than 13 renal ciliopathy genes13Halbritter J. Diaz K. Chaki M. et al.High-throughput mutation analysis in patients with a nephronophthisis-associated ciliopathy applying multiplexed barcoded array-based PCR amplification and next-generation sequencing.J Med Genet. 2012; 49: 756-767Crossref PubMed Scopus (98) Google Scholar, 14Halbritter J. Porath J.D. Diaz K.A. et al.Identification of 99 novel mutations in a worldwide cohort of 1,056 patients with a nephronophthisis-related ciliopathy.Hum Genet. 2013; 132: 865-884Crossref PubMed Scopus (165) Google Scholar at a time. As we identified the disease-causing mutation in ∼2/3 (50/79) of individuals with increased echogenicity on RUS, our data show that WES is an efficient tool for molecular diagnostics in this patient cohort. WES provides affected individuals and their families with an unequivocal diagnosis that avoids unnecessary diagnostic and therapeutic interventions. WES is a noninvasive diagnostic tool with little risk, and, recently, the cost has dropped to a level that makes it accessible to a broader group of affected individuals in a clinical rather than a research setting. Considering the genetic heterogeneity of NPHP-RC, WES is the only available technique that can reliably establish the correct molecular diagnosis. The use of targeted sequencing panels as utilized for other disease entities in clinical genetics is technically not feasible and/or not economical for such a large number of potentially causative genes. However, WES yields a large number of variants from normal reference sequence, most of which are functionally irrelevant. Therefore, an efficient strategy for variant filtering is indispensable for the work with WES data. In addition, variant calling should follow an indication-driven strategy: After thorough clinical evaluation, a subset of potentially causative genes should be defined prior to WES, and the evaluation should primarily focus on variants in these genes. For example, in a patient with increased echogenicity and/or ≥2 cysts on renal ultrasound, primarily variants in the ∼90 monogenic causes of renal ciliopathies, and genes that can phenocopy NPHP-RC, should be taken into consideration. In our cohort, WES molecularly confirmed the clinically suspected diagnosis of NPHP-RC in 64% of cases. However, 18/50 individuals (36%) harbored a mutation in a gene that causes a monogenic kidney disease different from NPHP-RC. As already described in a smaller cohort,1Gee H.Y. Otto E.A. Hurd T.W. et al.Whole-exome resequencing distinguishes cystic kidney diseases from phenocopies in renal ciliopathies.Kidney Int. 2014; 85: 880-887Abstract Full Text Full Text PDF PubMed Scopus (54) Google Scholar the majority of these cases (8/18) were diagnosed as renal tubulopathies. In addition, we identified four individuals with Alport syndrome, three individuals with mutations in CAKUT genes, two individuals with ARPKD, and one individual with autoimmune nephropathy in the context of APECED syndrome. In 10 of these 18 cases, the molecular diagnosis differed from the initial clinical diagnosis. We therefore suggest that WES helps distinguish between NPHP-RC and other kidney diseases that phenocopy the presentation of NPHP-RC on renal ultrasound. This can be particularly advantageous in very early stages of disease progression in which additional characteristic symptoms might not yet be present, as well as for very rare genetic syndromes. This observation underlines an additional benefit of WES: The genetic diagnostic is not restricted to a limited subset of monogenic diseases that were already suspected based on the clinical presentation, but a broader spectrum of monogenic causes can be taken into consideration. In addition, the very high rate of detecting a causative mutation in genes that are known to be mutated in renal disorders with increased echogenicity confirms that increased renal echogenicity, even if performed in numerous centers worldwide, is a reproducible and reliable criterion for noninvasive diagnostic in pediatric kidney disease. Interestingly, the percentage of molecularly diagnosed individuals was not considerably different in consanguineous families as compared with outbred sibling cases. We therefore suggest that the relevance of our findings reaches beyond consanguineous cases. As we and others have shown10Chaki M. Airik R. Ghosh Amiya K. et al.Exome capture reveals ZNF423 and CEP164 mutations, linking renal ciliopathies to DNA damage response signaling.Cell. 2012; 150: 533-548Abstract Full Text Full Text PDF PubMed Scopus (260) Google Scholar, 12Schueler M. Braun D.A. Chandrasekar G. et al.DCDC2 mutations cause a renal-hepatic ciliopathy by disrupting Wnt signaling.Am J Hum Genet. 2015; 96: 81-92Abstract Full Text Full Text PDF PubMed Scopus (68) Google Scholar, 17Zariwala M.A. Gee H.Y. Kurkowiak M. et al.ZMYND10 is mutated in primary ciliary dyskinesia and interacts with LRRC6.Am J Hum Genet. 2013; 93: 336-345Abstract Full Text Full Text PDF PubMed Scopus (143) Google Scholar combining WES and homozygosity mapping in consanguineous families represents a powerful tool for the identification of novel human disease genes. We applied this technique in five families in whom mutations in known human disease genes had been excluded, and we identified RBM48, FAM168B, PIAS1, INCENP, and RCOR1 as novel candidate genes for renal ciliopathies. So far, mutation analysis has not yielded additional families with mutations in these genes. However, this rarity is not unexpected as for the majority of recently identified NPHP-RC genes less than 10 families with mutations have been described worldwide.9Otto E.A. Trapp M.L. Schultheiss U.T. et al.NEK8 mutations affect ciliary and centrosomal localization and may cause nephronophthisis.J Am Soc Nephrol. 2008; 19: 587-592Crossref PubMed Scopus (147) Google Scholar, 10Chaki M. Airik R. Ghosh Amiya K. et al.Exome capture reveals ZNF423 and CEP164 mutations, linking renal ciliopathies to DNA damage response signaling.Cell. 2012; 150: 533-548Abstract Full Text Full Text PDF PubMed Scopus (260) Google Scholar, 11Otto E.A. Hurd T.W. Airik R. et al.Candidate exome capture identifies mutation of SDCCAG8 as the cause of a retinal-renal ciliopathy.Nat Genet. 2010; 42: 840-850Crossref PubMed Scopus (254) Google Scholar, 12Schueler M. Braun D.A. Chandrasekar G. et al.DCDC2 mutations cause a renal-hepatic ciliopathy by disrupting Wnt signaling.Am J Hum Genet. 2015; 96: 81-92Abstract Full Text Full Text PDF PubMed Scopus (68) Google Scholar, 18Chaki M. Hoefele J. Allen S.J. et al.Genotyp
Referência(s)